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We thank J. Peters, L. Zhang, J. Rittle, C. Morrison, B. Wenke, and
K. Dörner for informative discussions. This work was supported by
NIH grant GM45162 (D.C.R.), Deutsche Forschungsgemeinschaft
grants EI-520/7 and RTG 1976, and the European Research Council
N-ABLE project (O.E.). We gratefully acknowledge the Gordon
and Betty Moore Foundation, the Beckman Institute, and the
Sanofi-Aventis Bioengineering Research Program at Caltech for
their generous support of the Molecular Observatory at Caltech
and the staff at Beamline 12–2, Stanford Synchrotron Radiation
Lightsource (SSRL), for their assistance with data collection.
SSRL is operated for the U.S. Department of Energy and supported
by its Office of Biological and Environmental Research and by
the NIH: National Institute of General Medical Sciences (P41GM103393)
and the National Center for Research Resources (P41RR001209).
We thank the Center for Environmental Microbial Interactions for its
support of microbiology research at Caltech. Coordinates and
structure factors have been deposited in the Protein Data Bank of
the Research Collaboratory for Structural Bioinformatics, with IDs
4TKV (Av1-CO) and 4TKU (Av1 reactivated).
Materials and Methods
Figs. S1 and S2
Tables S1 and S2
29 May 2014; accepted 31 July 2014
Immune dysregulation in human
subjects with heterozygous germline
mutations in CTLA4
Hye Sun Kuehn,1 Weiming Ouyang,2 Bernice Lo,3,4 Elissa K. Deenick,5,6 Julie E. Niemela,1
Danielle T. Avery,5 Jean-Nicolas Schickel,7 Dat Q. Tran,8 Jennifer Stoddard,1 Yu Zhang,4,9
David M. Frucht,2 Bogdan Dumitriu,10 Phillip Scheinberg,10 Les R. Folio,11 Cathleen A. Frein,12
Susan Price,3,4 Christopher Koh,13 Theo Heller,13 Christine M. Seroogy,14 Anna Huttenlocher,14,15
V. Koneti Rao,3,4 Helen C. Su,4,9 David Kleiner,16 Luigi D. Notarangelo,17 Yajesh Rampertaap,18
Kenneth N. Olivier,18 Joshua McElwee,19 Jason Hughes,19 Stefania Pittaluga,16 Joao B. Oliveira,20
Eric Meffre,7 Thomas A. Fleisher,1† Steven M. Holland,4,18 Michael J. Lenardo,3,4†
Stuart G. Tangye,5,6 Gulbu Uzel18†
Cytotoxic T lymphocyte antigen–4 (CTLA-4) is an inhibitory receptor found on immune
cells. The consequences of mutations in CTLA4 in humans are unknown. We identified
germline heterozygous mutations in CTLA4 in subjects with severe immune
dysregulation from four unrelated families. Whereas Ctla4 heterozygous mice have
no obvious phenotype, human CTLA4 haploinsufficiency caused dysregulation of FoxP3+
regulatory T (Treg) cells, hyperactivation of effector T cells, and lymphocytic infiltration
of target organs. Patients also exhibited progressive loss of circulating B cells, associated
with an increase of predominantly autoreactive CD21lo B cells and accumulation of B cells
in nonlymphoid organs. Inherited human CTLA4 haploinsufficiency demonstrates a critical
quantitative role for CTLA-4 in governing T and B lymphocyte homeostasis.
Immune tolerance is controlled by multiple mechanisms (1,2), includingregulatory T(Treg) cells (3–5) and inhibitory receptors (6, 7). Treg cells constitutively express the inhibitory re- ceptor CTLA-4, which confers suppressive
functions (8, 9). CTLA-4, also known as CD152,
is also expressed by activated T cells and, upon
ligation, inhibits their proliferation (10). Homozygous deficiency of Ctla4 in mice causes fatal
multiorgan lymphocytic infiltration and destruction (11–13); hence, CTLA-4 functions at a key
“checkpoint” in immune tolerance. CTLA-4–
immunoglobulin (Ig) fusion protein and neutralizing
CTLA-4 antibody are used to modulate immunity in
autoimmune and cancer patients (14, 15), respectively. Studies have given conflicting results regarding the association of CTLA4 single-nucleotide
variants (SNVs) with organ-specific autoimmunity
(16). The consequences of genetic CTLA-4 deficiency in humans are unknown.
Our index patient—a 22-year-old female (A.II.1)—
developed brain, gastrointestinal (GI), and lung
lymphocytic infiltrates, autoimmune thrombocytopenia, and hypogammaglobulinemia in early
childhood (Fig. 1A and table S1). Her 43-year-old
father (A.I.1) manifested lung and GI infiltrates,
hypogammaglobulinemia, and clonally expanded
gd-CD8+ T cells infiltrating and suppressing the
bone marrow (fig. S1A). Four additional cases
from three unrelated families (families B, C,
and D) (fig. S1 and table S1) were identified
among a cohort of 23 patients with autoimmune
cytopenias, hypogammaglobulinemia, CD4 T cell
lymphopenia, and lymphocytic infiltration of nonlymphoid organs. Patient B.I.1, previously diagnosed with common variable immunodeficiency
1Department of Laboratory Medicine, Clinical Center, National
Institutes of Health, Bethesda, MD 20892, USA. 2Laboratory
of Cell Biology, Division of Monoclonal Antibodies, Office of
Biotechnology Products, Center for Drug Evaluation and
Research, U.S. Food and Drug Administration, Bethesda,
MD 20892, USA. 3Molecular Development of the Immune
System Section, Laboratory of Immunology, National
Institute of Allergy and Infectious Diseases, Bethesda, MD
20892, USA. 4NIAID Clinical Genomics Program, National
Institute of Allergy and Infectious Diseases, Bethesda, MD
20892, USA. 5Immunology and Immunodeficiency Group,
Immunology Division, Garvan Institute of Medical Research,
Sydney, NSW 2010, Australia. 6St. Vincent’s Clinical School
Faculty of Medicine, University of New South Wales, Sydney,
NSW 2010, Australia. 7Department of Immunobiology, Yale
University School of Medicine, New Haven, CT 06511, USA.
8Department of Pediatrics, University of Texas Medical
School, Houston, TX 77030, USA. 9Immunological Diseases
Unit, Laboratory of Host Defenses, National Institute of
Allergy and Infectious Diseases, Bethesda, MD 20892, USA.
10Hematology Branch, National Heart, Lung and Blood
Institute, Bethesda, MD 20892, USA. 11Radiology and
Imaging and Sciences, Clinical Center, National Institutes of
Health, Bethesda, MD 20892, USA. 12Clinical Research
Directorate, Clinical Monitoring Research Program, Leidos
Biomedical Research Inc., Frederick National Laboratory
for Cancer Research, Frederick, MD 21702, USA. 13Liver
Diseases Branch, National Institute of Diabetes and
Digestive and Kidney Diseases, Bethesda, MD 20892, USA.
14Department of Pediatrics, University of Wisconsin, Madison,
WI 53706, USA. 15Department of Medical Microbiology and
Immunology, University of Wisconsin, Madison, WI 53706,
USA. 16Laboratory of Pathology, National Cancer Institute,
Bethesda, MD 20892, USA. 17Division of Immunology and
Manton Center for Orphan Disease Research, Children’s
Hospital, Harvard Medical School, Boston, MA 10217, USA.
18Laboratory of Clinical Infectious Diseases, National Institute
of Allergy and Infectious Diseases, Bethesda, MD 20892,
USA. 19Merck Research Laboratories, Merck & Co., Boston,
MA 02130, USA. 20Instituto de Medicina Integral Prof.
Fernando Figueira–IMIP, 50070 Recife-PE, Brazil.
*These authors contributed equally to this work.
†Corresponding author. E-mail: firstname.lastname@example.org (T.A.F.);
email@example.com (M.J.L.); firstname.lastname@example.org (G.U.)